Genetic and epigenetic screens in primary human T cells link candidate causal autoimmune variants to T cell networks [gRNA]

Genetic variants associated with autoimmune diseases are highly enriched within putative cis-regulatory regions of CD4+ T cells, suggesting that they alter disease risk via changes in gene regulation. However, very few genetic variants have been shown to affect T cell gene expression or function. Here, we tested >18,000 autoimmune disease-associated variants for allele-specific expression using massively parallel reporter assays in primary human CD4+ T cells. We find 545 variants that modulate expression in an allele-specific manner (emVars). Primary T cell emVars greatly enrich for probable causal variants, are mediated by common upstream pathways, and their putative target genes are highly enriched within a lymphocyte activation network. Using bulk and single-cell CRISPR-interference screens, we confirm that emVar-containing T cell cis-regulatory elements modulate both known and previously unappreciated target genes that regulate T cell proliferation, providing plausible mechanisms by which these variants alter autoimmune disease risk. Overall design: We created gRNA libraries targeting on average 14 gRNAs to each variant CRE (within 100 bp of each variant), 120 positive control gRNAs targeting known regulators of T cell proliferation and effector function, and 992 non-targeting control gRNAs We activated primary human T cells and delivered dCas9- ZIM3 and the gRNA library using lentivirus infection. We harvested half of the transduced cells directly post-puromycin treatment (day 2) and the remainder of the cells after 21 days of proliferation (day 21).